Breast cancer is the most common cancer of women in the United States. Despite recent improvements in conventional treatments, advanced breast cancer still has an extremely poor prognosis, resulting in more than 45,000 deaths each year in the US alone. The development of new therapeutic modalities is therefore of great importance. The long-term goal of the research outlined in this proposal is to develop an effective and safe virotherapy for metastatic breast cancer. The central hypothesis is that the antitumor activity of an oncolytic herpes simplex virus (HSV) can be significantly enhanced by incorporating a cell-membrane fusion function into the virus, which will produce syncytia formation in the tumor, thereby directly enhancing the destructive power of the virus and promoting its intratumor spread even in the presence of host's antiviral immunity. The second hypothesize is that the unique mechanism of tumor destruction in vivo by the fusogenic oncolytic HSV can induce strong antitumor immune responses, which can further facilitate tumor eradication. Specific Aim 1 seeks to determine if a doubly fusogenic oncolytic HSV, which was constructed by incorporating two independent cell-membrane fusion mechanisms into the virus, can provide effective and long-term therapy to distant organ metastases of breast cancer. The safety of this virus will also be fully assessed in this aim. Specific Aim 2 sets to explore the ability of tumor destruction by the fusogenic oncolytic HSV to induce antitumor immunity. First, the antitumor effect and the accompanying antitumor immunity induced by fusogenic and nonfusogenic oncolytic HSVs will be directly compared in a murine mammary tumor model. Then antibody depletion of immune cells (e.g., CD4+ and CD8+ T cells) will be used to determine if the tumor-specific immune response directly contributes to tumor eradication and which immune cells are responsible for the antitumor immunity. Experiments will also be conducted to dissect the mechanism of enhancement of antitumor immunity by fusogenic oncolytic HSVs. In Aim 3, the influences of pre-existing antiviral immunity on spread and antitumor effect of fusogenic and non-fusogenic HSVs in metastastic breast will be examined. Then experiments will be conducted to determine if systemic delivery of oncolytic HSV through liposome-formulation of viral DNA or through cell-carriers can evade host's antiviral immunity. The proposed studies will establish a strong preclinical rationale for using the fusogenic oncolytic HSV to treat metastatic breast cancer and will serve as the necessary foundation for a human clinical trial. Finally, if successful in breast cancer, this therapeutic strategy may be applicable to other solid tumors.